title: Involvement of boxA nucleotides in the formation of a stable ribonucleoprotein complex containing the bacteriophage lambda N protein.
abstract: The association of the transcriptional antitermination protein N of bacteriophage lambda with Escherichia coli RNA polymerase depends on nut site RNA (boxA + boxB) in the nascent transcript and the host protein, NusA. This ribonucleoprotein complex can transcribe through Rho-dependent and intrinsic termination sites located up to several hundred base pairs downstream of nut. For antitermination to occur farther downstream, this core antitermination complex must be stabilized by the host proteins NusB, NusG, and ribosomal protein S10. Here, we show that the assembly of NusB, NusG, and S10 onto the core complex involves nucleotides 2-7 of lambda boxA (CGCUCUUACACA) and is a fully cooperative process that depends on the presence of all three proteins. This assembly of NusB, NusG, and S10 also requires the carboxyl-terminal region (amino acids 73-107) of N, which interacts directly with RNA polymerase. NusB and S10 assemble in the absence of NusG when lambda boxA is altered at nucleotides 8 and 9 to create a consensus version of boxA (CGCUCUUUAACA). These experiments suggest that multiple protein-protein and protein-RNA interactions are required to convert a core antitermination complex into a complete complex.
authors.affiliation: Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario M5G 1L6, Canada.
authors.forename: J
authors.initials: J
authors.lastname: Mogridge
authors.forename: T F
authors.initials: TF
authors.lastname: Mah
authors.forename: J
authors.initials: J
authors.lastname: Greenblatt
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D010582:Bacteriophage lambda; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D029968:Escherichia coli Proteins; D016296:Mutagenesis / Q000235:genetics; D010445:Peptide Elongation Factors / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D016601:RNA-Binding Proteins / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism*; D017384:Sequence Deletion / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics; D035602:Transcriptional Elongation Factors; D014764:Viral Proteins / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C059417:N protein, Bacteriophage lambda; C074952:NusB protein, E coli; C070291:NusG protein, E coli; D010445:Peptide Elongation Factors; D012329:RNA, Bacterial; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D014157:Transcription Factors; D035602:Transcriptional Elongation Factors; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C068727:nusA protein, E coli; D012321:DNA-Directed RNA Polymerases